The expression of osteopontin (OPN) and one of its ligands, CD44, was studied in the spinal cord of rats with experimental autoimmune encephalomyelitis (EAE). Western blot analysis showed that osteopontin significantly increased at the early and peak stage of EAE and slightly declined thereafter. Osteopontin was constitutively expressed in some astrocytes adjacent to pia mater and neurons in normal rats, and was shown to be increased in the same cells and also in some inflammatory cells including macrophages at the early and peak stage of EAE. CD44, a ligand for osteopontin, was constitutively expressed in astrocytes in normal and control spinal cords and was also expressed in inflammatory cells, as well as increased expression in astrocytes in EAE. These findings suggest that inflammatory cells as well as reactive astrocytes are major sources of osteopontin in rat EAE, and osteopontin may interact with its ligand CD44 on astrocytes and inflammatory cells in EAE, possibly mediating autoimmune central nervous system (CNS) diseases in rats.